Methods for Data-Dependent Mass Spectrometry of Mixed Biomolecular Analytes

A method for mass spectral analysis of a sample containing a plurality of intact protein molecule species, said method including repeatedly introducing a respective portion of the sample into an ionization source of a mass spectrometer;

• and using the ionization source to generate, from each sample portion, a population of multi-protonated ions that comprises a plurality of ion species generated from each respective intact protein molecule species of said each portion, each ion species comprising a respective charge state, z, and mass-to-charge (m/z) ratio, the method comprising:
• (a) mass analyzing a plurality of ions generated from a first one of the sample portions, thereby generating a mass spectrum thereof, wherein each ion species generated from a protein is represented in the mass spectrum by a respective m/z ratio and a corresponding respective intensity;
• (b) automatically recognizing, for each of two or more intact protein molecule species, a respective subset of the m/z ratios corresponding to a set of ion species generated from said each intact protein molecule species and automatically determining a respective charge state, z, for each m/z ratio of each subset;
• (c) determining a respective molecular weight of each of the two or more intact protein molecule species from the m/z ratios and determined charge states of the two or more recognized subsets of the m/z ratios;
• and
• (d) reporting information relating to each subset of the m/z ratios and information relating to the two or more molecular weights to a user in the form of a displayed graph comprising:
• a horizontal axis scaled in units of m/z ratio;
• a vertical axis scaled in units of molecular weight;
• and
• a plurality of points plotted on the graph, each point representing a respective ion species in the mass spectrum for which an m/z value is recognized, wherein a horizontal coordinate of each plotted point represents the recognized m/z value of the respective ion species and a vertical coordinate of each plotted point represents the determined molecular weight of the respective intact protein molecule species.